Abstract

The usefulness of a recombinant phage library depends on the ability to screen a large number of phage and identify the clone
that carries the DNA sequence of interest. This unit presents a protocol in which phage are allowed to multiply in host bacteria
in a thin layer of agarose on regular bacterial plates. When nitrocellulose is applied to the agarose, phage particles and
unpackaged DNA adsorb to the filter to produce a replica of the plate surface. If the agarose surface is not excessively wet,
there will be little spreading of the phage on the filter. Subsequent treatment of the filter with sodium hydroxide destroys
the phage particles and denatures the phage DNA which then binds to the nitrocellulose. Neutralization of the filters is required
to maintain the integrity of the nitrocellulose. Hybridization of these filters to a DNA or RNA probe will identify the location
of the phage plaque of interest, which can then be recovered from the plate.